The “dilution effect” in stallion sperm

Dilution of semen to less than 20 × 106 sperm/mL has been reported to decrease sperm quality in multiple species, a phenomenon known as the semen “dilution effect.” Critical evaluation of stallion semen diluted to these concentrations, however, has not been reported. This study evaluated sperm motion characteristics (percent total motility [TMOT], percent progressive motility [PMOT], curvilinear velocity [μm/s], and percent straightness) and plasma membrane integrity (percent plasma membrane intact [PMI]) in semen samples diluted to 2.5 × 106 sperm/mL with the addition of 0%, 7.5%, or 25% seminal plasma (groups T-2.5/0, T-2.5/7.5, and T-2.5/25, respectively), or after simple dilution to 30 × 106 sperm/mL (group T-30), or simple dilution to a ratio of 3:1 (extender:semen; group T-3:1SD). Evaluations were performed immediately after semen collection (T0), and after 24 and 48 hours of cooled storage (T24 and T48, respectively). The PMI and TMOT were the highest in group T-3:1SD at T0. At T24, the PMI in groups T-30, T3:1SD and T3:1/30, and T-2.5/0 were higher than that in the other groups (P < 0.05), whereas TMOT in group T-3:1SD was higher (P < 0.05) than that in all other groups except T-30. By T48, no difference was detected for PMI among groups T-3:1SD, T-30, and T-2.5/0; for TMOT among groups T-3:1SD, T-30, and T-2.5/0, and T-2.5/7.5 (P > 0.05), whereas PMOT was the highest in groups T-2.5/0 and T-2.5/7.5 (P < 0.05). These findings revealed that treatments in which semen was diluted to a concentration of 2.5 × 106 sperm/mL had lower initial PMI, TMOT, and PMOT, but semen quality did not decline after 24 and 48 hours of cooled storage. In this study, TMOT and PMI in dilute semen were less than those in more concentrated semen at T0. This effect, while significant, was small and less apparent after cooled storage.